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Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restriction (Only characterization instead of identification of the metabolites. Analytical purity not reported).

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
The pharmacokinetics of inhaled chlorobenzene in the rat
Author:
Sullivan, T.M. et al.
Year:
1983
Bibliographic source:
Toxicol. Appl. Pharmacol. 71:194 - 203
Reference Type:
secondary source
Title:
Environmental toxicology: organic pollutants. Ellis Horwood limited
Author:
Fawell J.K., Hunt S. 1988. Environmental toxicology: organic pollutants.
Year:
1988
Bibliographic source:
Ellis Horwood limited.Publishers-Chichester. Halsted Press: a division of John wiley & Sons. New York-Chichester-Brisbane-Toronto

Materials and methods

Objective of study:
distribution
excretion
metabolism
Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
Remarks:
Only characterization instead of identification of the metabolites. Analytical purity not reported
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Chloro[U-14C]benzene
- Analytical purity: no data
- Radiochemical purity (if radiolabelling): 98%
- Specific activity (if radiolabelling): 1.12 µCi/µmol
Radiolabelling:
yes
Remarks:
Chloro[U-14C]benzene

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Laboratory Supply Co., Indianapolis, Ind., USA
- Age at study initiation: 58-65 days old
- Weight at study initiation: 160-210 g
- Housing: Roth type metabolism cage after exposure
- Individual metabolism cages: yes
- Diet (e.g. ad libitum): lab chow ad libitum
- Water (e.g. ad libitum): tap water


Administration / exposure

Route of administration:
inhalation: vapour
Vehicle:
unchanged (no vehicle)
Details on exposure:
TYPE OF INHALATION EXPOSURE: whole body


GENERATION OF TEST ATMOSPHERE / CHAMPER DESCRIPTION
- Exposure apparatus: 40-liter glass chromatography jar
- Source and rate of air: [14C]-chlorobenzene vapor was generated by passing a portion of the total air flow though a heated flask containing glass beads. The[14C]-chlorobenzene was infused through a septum immediately upstream from the flask using a gas-tight syringe ans variable speed syringe pump. The vapor passed through a mixing chamber before re-entering the main air flow. Total air flow was 14 liters/min.
- Concentration of test material in vehicle (if applicable): 100 ppm, 400 ppm, and 700 ppm

Duration and frequency of treatment / exposure:
8 hours per day for either 1 day or 5 days
Doses / concentrations
Remarks:
Doses / Concentrations:
Single exposure: 100, 400, 700 ppm. Repeated exposure :400 ppm
No. of animals per sex per dose:
100, 400, and 700 ppm group for 8 hours: 15 animals
Multiple exposure regimen : 6 animal each group for 5 days
3 animals for the first 4 days of the multiple exposure regimen. 6 rats exposed only the fifth day of every exposure block (single exposure regimen)
12 rats were exposed in a 1-day repeat study at 400 ppm
Control animals:
no
Details on study design:
- Dose selection rationale: the authors intended to examine the dose dependency of parameters indicative of the toxicity process and effect of repeate exposure. Moreover 400 ppm concentration was choosed to account for the statistical validity of the blocking effect.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled (delete / add / specify): urine, blood, epididymal fat, perinatal fat, kidneys, lungs, liver





Statistics:
The design of the treatment procedure allowed only one exposure concentration to be used in any given exposure block. The significance of the blocking factor was estimated in a separate analysis of variance (ANOVA) using the two groups of singly exposed rats exposed to 400 ppm. The magnitude of all other treatment effects was estimated by performing ANOVA on the data from the 100, 700, and the first 400 ppm exposure blocks, excluding the data from the repeated 400 ppm block. Comparison between treatment groups were based on the t statistic.

Results and discussion

Preliminary studies:
No significant fecal excretion of chlorobenzene or its metabolites occurs in rats (Williams et al., 1959). Preliminary studies indicated that [14C]-labeled material excreted in the urine and expired air after 48 hr postexposure amounted to no more that 2% of the total excreted from 0 to 48 hr.

Toxicokinetic / pharmacokinetic studies

Details on distribution in tissues:
There was no evidence of of accumulation in the rats sacrificed immediately after exposure, except in kidney (single exposure 0hr vs multiple exposure 0 -hr). There was a tendency at the 48 hrs after exposure for mutiply exposed rats to exhibit higher tissue burdens than rats exposed only once. The [14C]chlorobenzene of adipose tissue increased about 8 to 10 fold when the concentration was increased from 100 to 400 ppm and about 3 to 5 fold from 400 to 700 ppm.
Transfer into organs
Test no.:
#1
Transfer type:
other: inhaltion/lungs/ blood / target organs
Observation:
distinct transfer
Details on excretion:
14 [C]-Chlorobenzene and/or metabolites excreted in the urine and expired air are shown in table 1. Significant differences between multiply and singly exposed rats were observed at all the three concentrations and among all three concentations for both exposure regimens. The percentage of material excreted via exhalation increased with increasing exposure concentration and was decreased among multiply exposed rats compared to those singly exposed. There was no effect of exposure regimen in the total micromoles of chlorobenzene equivalents excreted in the urine (see Table 1) . There was a significant difference in the amount of 14 C-chlorobenzene expired between the two 400 ppm, singly exposed groups of rats, with the rats of the repeat study expiring more than rats in the initial group.
Toxicokinetic parametersopen allclose all
Test no.:
#1
Toxicokinetic parameters:
half-life 1st: 0.81 hr. Apparent half-lives of respiratory elimination following 100 ppm single inhalation exposure of rats to 14C-Chlorobenzene. Fast phase
Test no.:
#1
Toxicokinetic parameters:
half-life 2nd: 8.77 hr. Apparent half-lives of respiratory elimination following 100 ppm single inhalation exposure of rats to 14C-Chlorobenzene. Slow phase
Test no.:
#1
Toxicokinetic parameters:
half-life 1st: 0.60 hr. Apparent half-lives of respiratory elimination following 100 ppm single inhalation exposure of rats to 14C-Chlorobenzene. Fast phase
Test no.:
#1
Toxicokinetic parameters:
half-life 2nd: 9.24 hr. Apparent half-lives of respiratory elimination following 100 ppm multiple inhalation exposure of rats to 14C-Chlorobenzene. Slow phase (Data at the 12-, 24-, 36-, and 48- hour sampling times described the terminal portion of the respiratory excr
Test no.:
#2
Toxicokinetic parameters:
half-life 1st: 1.13 hr. Apparent half-lives of respiratory elimination following 400 ppm single inhalation exposure of rats to 14C-Chlorobenzene. Fast phase ( The more rapid phase of respiratory elimination).
Test no.:
#2
Toxicokinetic parameters:
half-life 2nd: 7.62 hr . Apparent half-lives of respiratory elimination following 400 ppm single inhalation exposure of rats to[14C-Chlorobenzene. Slow phase
Test no.:
#2
Toxicokinetic parameters:
half-life 1st: 0.83 hr. Apparent half-lives of respiratory elimination following 400 ppm multiple inhalation exposure of rats to [14C]-chlorobenzene. Fast phase
Test no.:
#2
Toxicokinetic parameters:
half-life 2nd: 8.56 hr. Apparent half-lives of respiratory elimination following 400 ppm multipleinhalation exposure of rats to [14C]-chlorobenzene. Slow phase
Test no.:
#3
Toxicokinetic parameters:
half-life 1st: 1.19 hr. Apparent half-lives of respiratory elimination following 400 ppm single inhalation exposure of rats to [14C]-chlorobenzene. Fast phase
Test no.:
#3
Toxicokinetic parameters:
half-life 3rd: 7.70 hr. Apparent half-lives of respiratory elimination following 400 ppm single inhalation exposure of rats to [14C]-chlorobenzene. Slow phase
Test no.:
#4
Toxicokinetic parameters:
half-life 1st: 3.65 hr. Apparent half-lives of respiratory elimination following 700 ppm single inhalation exposure of rats to [14C]-chlorobenzene. Fast phase
Test no.:
#4
Toxicokinetic parameters:
half-life 2nd: 5.73 hr. Apparent half-lives of respiratory elimination following 7400 ppm single inhalation exposure of rats to [14C]-chlorobenzene. Slow phase
Test no.:
#4
Toxicokinetic parameters:
half-life 1st: 1.62 hr. Apparent half-lives of respiratory elimination following 700 ppm multiple inhalation exposure of rats to [14C]-chlorobenzene. Fast phase
Test no.:
#4
Toxicokinetic parameters:
half-life 2nd: 6.60 hr. Apparent half-lives of respiratory elimination following 700 ppm multiple inhalation exposure of rats to [14C]-chlorobenzene. Slow phase

Metabolite characterisation studies

Metabolites identified:
no
Details on metabolites:
The metabolite were only characterised and seperated into four categories: 1) the initial category contained metabolites extracted into ether from raw, untreated urine; 2) the G/S category contained residues which had been excreted as glucuronid (G) or sulfate (S) conjugates; 3) mercapturic acid category; 4) unexctracted included all metabolites not extracted from the urine. Differences between singly and multiply exposed groups were not significant.

Any other information on results incl. tables

With regard to metabolites, the 3 concentrations (100, 400, and 700 ppm) resulted in signifcantly different mercapturic acid percentages among the singly exposed groups (p< 0.05). In the multiply exposed groups, the difference between the 400- and the 700 ppm results was not signifcant, although both were significantly decreased from the 100 ppm results. When the exposure concentration was increased from 100 to 700 ppm, the mercapturic acid percentage of the total was reduced by 27 % for the singly exposed group and 24 % for the multiply exposed group.

Table 4 presents the apparent half-lives (t1/2) calculated from regression models obtained by combining all rats in each treatment group. The exposure regimen effect was significant only at the 700 ppm concentration, with multiply exposed rats yielding a shorter fast phase t1/2 than singly exposed rats. There was no signifcant effect of concentration on respiratory elimination between the 100- and 400 ppm groups. However, the fast phase half-lives of the 700 ppm rats were longer than those observed at lower exposure concentrations.

Applicant's summary and conclusion

Executive summary:

Sullivan et al., 1983

Male Sprague-Dawly rats were exposed for 8 hr/day to 100, 400, or 700 ppm of [14C]-chlorobenzene vapor for either 1 day or 5 days. The study was conducted according to OECD guideline 417 with deviations (Only characterization instead of identification of the metabolites. Analytical purity not reported). 6 rats of each group were exposed for 5 consecutive days (multiple exposure regimen). Another 3 rats were exposed for the first 4 days of the multiple exposure regimen, but were sacrificed after 16 hr their final exposure. The other 6 rats were exposed only on the fifth day on each exposure block (single exposure regimen). Another group of 12 rats was exposed in a 1-day repeated study at 400 ppm to account for the statistical signifcance of blocking effect.

Immediataley after the fifth day of exposure, 6 of the 12 remaining rats (3 singly exposed and 3 multiply exposed) were placed in metabolism cages for collection of urine and expired material. The other 6 rats were sacrificed 48 hr after exposure for assessment of remaining tissue burdens.

There was no evidence of accumulation in the rats sacrificed immediately after exposure, except in kidney (single exposure 0hr vs multiple exposure 0 -hr). There was a tendency at the 48 hrs after exposure, for mutiply exposed rats to exhibit higher tissue burdens than rats exposed only once. The [14C]chlorobenzene of adipose tissue increased about 8 to 10 fold when the concentration was increased from 100 to 400 ppm and about 3 to 5 fold from 400 to 700 ppm.

Respiratory elimination of [14C]chlorobenzene also increased in a dose-dependent way. There are no differences due to dosing regimen in the amount of material excreted in the urine. There was a significant difference in the amount of 14 C-chlorobenzene expired between the two 400 ppm, singly exposed groups of rats, with the rats of the repeat study expiring more than rats in the initial group.

The exposure regimen effect was significant only at the 700 ppm concentration, with multiply exposed rats yielding a shorter fast phase t1/2 than singly exposed rats. There was no signifcant effect of concentration on respiratory elimination between the 100- and 400 ppm groups. However, the fast phase half-lives of the 700 ppm rats were longer than those observed at lower exposure concentrations.

Analysis of treatment effects in the metabolite profile was based on the percentage of mercapturic acid, since this is the paramenter most relavant to toxicity. Differences between singly and multiply exposed groups were not significant. The three concentrations resulted in significantly different mercapturic acid percentages among the singly exposed groups. In the multiply exposed groups, the difference between the 400 ppm and the 700 ppm results was not significant, although were significantly decreased from 100 ppm results. When the exposure concentration was increased from 100 to 700 pppm, the mercapturic acid percentage of the total was reduced by 27 % for singly exposed group and 24% for the multiply exposed group.

The dose-dependent changes are postulated to be due to saturation of the metabolic elimination of chlorobenzene. The effect of multiple exposure is apparently some stimulation of metabolism.